Protein phosphatase 2A regulates deoxycytidine kinase activity <i>via</i> Ser‐74 dephosphorylation

Amsailale, Rachid; Beyaert, Maxime; Smal, Caroline; Janssens, Veerle; Neste, Éric Van Den; Bontemps, Françoise

doi:10.1016/j.febslet.2014.01.016

Cited by 6 publications

(7 citation statements)

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“…These data are in line with our observations of reduced dCK phosphorylation/activation in the clofarabine resistant p.R183W Aα expressing cells. dCK is activated by phosphorylation on Ser74, a site reported to be dephosphorylated by PP2A [31,37]. Accordingly, we showed that combination of OA with clofarabine was able to reactivate dCK and sensitize the cells to clofarabine, with the most pronounced effect on the p.R183W Aα expressing cells, and thus, suggesting that an unexpected PP2A gain-of-function was responsible for the observed phenotype in these cells.…”

Section: Discussionmentioning

confidence: 60%

The PPP2R1A cancer hotspot mutant p.R183W increases clofarabine resistance in uterine serous carcinoma cells by a gain-of-function mechanism

Remmerie,

Dok,

Wang

et al. 2024

Preprint

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Purpose: Uterine serous carcinoma (USC) is generally associated with poor prognosis due to a high recurrence rate and frequent treatment resistance; hence, there is a need for improved therapeutic strategies. Molecular analysis of USC identified several molecular markers, useful to improve current treatments or identify new druggable targets. PPP2R1A, encoding the Aα subunit of the tumor suppressive Ser/Thr phosphatase PP2A, is mutated in up to 40% of USC. Here, we investigated the effect of the p.R183W PPP2R1A hotspot variant on treatment response to the nucleoside analogue clofarabine. Methods & Results: USC cells stably expressing p.R183W Aα showed increased resistance to clofarabine treatment in vitro and in vivo, corroborated by decreased clofarabine-induced apoptosis, G1 phase arrest, DNA-damage (γH2AX) and activation of ATM and Chk1/2 kinases. Phenotypic rescue by pharmacologic PP2A inhibition or dicer-substrate siRNA (dsiRNA)-mediated B56δ subunit knockdown supported a gain-of-function mechanism of Aα p.R183W, promoting dephosphorylation and inactivation of deoxycytidine kinase (dCK), the cellular enzyme responsible for the conversion of clofarabine into its bioactive form. Therapeutic assessment of related nucleoside analogues (gemcitabine, cladribine) revealed similar effects, but in a cell line-dependent manner. Expression of two other PPP2R1A USC mutants (p.P179R or p.S256F) did not affect clofarabine response in our cell models, arguing for mutant-specific effects on treatment outcome as well. Conclusions: While our results call for PPP2R1A mutant and context-dependent effects upon clofarabine/nucleoside analogue monotherapy, combining clofarabine with a pharmacologic PP2A inhibitor proved synergistically in all tested conditions, highlighting a new generally applicable strategy to improve treatment outcome in USC.

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Section: Discussionmentioning

confidence: 60%

The PPP2R1A cancer hotspot mutant p.R183W increases clofarabine resistance in uterine serous carcinoma cells by a gain-of-function mechanism

Remmerie,

Dok,

Wang

et al. 2024

Preprint

Self Cite

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“…27,40 dCK phosphorylation at serine 74 is reversed by protein phosphatase 2A, which negatively regulates dCK activity. 41 Moreover, using a mass spectrometry technique, dCK was found to exist in a complex that contains cyclin-dependent kinase 1 (Cdk1). After IR, Cdk1 interacts with dCK, and the activity of Ckd1 is inhibited by dCK both in vitro and in vivo, making dCK an important G2/M checkpoint regulator in response to DNA damage.…”

Section: Stimulates Ros Generation In Pancreatic Cancer Cells Resultingmentioning

confidence: 99%

“…dCK activation shifts dCK substrate specificity towards deoxycytidine, increases the intracellular dCTP pools and DNA repair activity, and contributes to chemotherapy and radiotherapy resistance . dCK phosphorylation at serine 74 is reversed by protein phosphatase 2A, which negatively regulates dCK activity . Moreover, using a mass spectrometry technique, dCK was found to exist in a complex that contains cyclin‐dependent kinase 1 (Cdk1).…”

Section: Discussionmentioning

confidence: 99%

dCK negatively regulates the NRF2/ARE axis and ROS production in pancreatic cancer

Qin

Xiang

et al. 2018

Cell Proliferation

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Objectives: Decreased deoxycytidine kinase (dCK) expression is a reported indicator of gemcitabine efficacy in pancreatic cancer, due to the impact of this kinase on gemcitabine metabolism. The transcription factor NF-E2 p45-related factor 2 (NRF2, also called Nfe2l2), a master regulator of redox homoeostasis, has been reported to tightly control the expression of numerous ROS-detoxification genes and participates in drug resistance. However, the contribution of dCK to the NRF2 signalling axis has seldom been discussed and needs investigation. Materials and methods:By overexpressing dCK in pancreatic cancer cells, we assessed the impact of dCK on NRF2 transcriptional activity. Furthermore, we measured the impact of dCK expression on the intracellular redox balance and reactive oxygen species (ROS) production. By utilizing immunohistochemical staining and tissues from pancreatic cancer patients, we assessed the correlation between dCK and NRF2 expression. Through proliferation and metastasis assays, we examined the impact of dCK expression on cell proliferation and metastasis.Results: dCK negatively regulates NRF2 transcriptional activity, leading to the decreased expression of ARE-driven antioxidant genes. In addition, dCK negatively regulates intracellular redox homoeostasis and ROS production. Negative correlations between dCK and NRF2 levels in pancreatic cancer cell lines and patient samples were observed. In vitro cell line studies suggested that dCK negatively regulated proliferation and metastasis. Conclusion:Decreased dCK expression promotes NRF2-driven antioxidant transcription, which further enhances gemcitabine treatment resistance, forming a feedback loop.This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

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“…For example, using gemcitabine treatment in tumours of different origin such as pancreas and lung, it was shown that resistance only is predicted by dCK activity and protein level, and not by dCK mRNA level [ 28 ], in agreement with our data. DCK enzymatic activity is controlled by phosphorylation at Ser-74 [ 29 ], and dephosphorylation decreases enzyme activity [ 30 ]. A more active mutant of dCK, with a 10,000-fold increased sensitivity to nucleoside analogues has been created and could be of interest for suicide gene approaches [ 10 , 31 ].…”

Section: Discussionmentioning

confidence: 99%

Bortezomib prevents cytarabine resistance in MCL, which is characterized by down-regulation of dCK and up-regulation of SPIB resulting in high NF-κB activity

et al. 2018

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BackgroundThe addition of high-dose cytarabine to the treatment of mantle cell lymphoma (MCL) has significantly prolonged survival of patients, but relapses are common and are normally associated with increased resistance. To elucidate the mechanisms responsible for cytarabine resistance, and to create a tool for drug discovery investigations, we established a unique and molecularly reproducible cytarabine resistant model from the Z138 MCL cell line.MethodsEffects of different substances on cytarabine-sensitive and resistant cells were evaluated by assessment of cell proliferation using [methyl-14C]-thymidine incorporation and molecular changes were investigated by protein and gene expression analyses.ResultsGene expression profiling revealed that major transcriptional changes occur during the initial phase of adaptation to cellular growth in cytarabine containing media, and only few key genes, including SPIB, are deregulated upon the later development of resistance. Resistance was shown to be mediated by down-regulation of the deoxycytidine kinase (dCK) protein, responsible for activation of nucleoside analogue prodrugs. This key event, emphasized by cross-resistance to other nucleoside analogues, did not only effect resistance but also levels of SPIB and NF-κB, as assessed through forced overexpression in resistant cells. Thus, for the first time we show that regulation of drug resistance through prevention of conversion of pro-drug into active drug are closely linked to increased proliferation and resistance to apoptosis in MCL. Using drug libraries, we identify several substances with growth reducing effect on cytarabine resistant cells. We further hypothesized that co-treatment with bortezomib could prevent resistance development. This was confirmed and show that the dCK levels are retained upon co-treatment, indicating a clinical use for bortezomib treatment in combination with cytarabine to avoid development of resistance. The possibility to predict cytarabine resistance in diagnostic samples was assessed, but analysis show that a majority of patients have moderate to high expression of dCK at diagnosis, corresponding well to the initial clinical response to cytarabine treatment.ConclusionWe show that cytarabine resistance potentially can be avoided or at least delayed through co-treatment with bortezomib, and that down-regulation of dCK and up-regulation of SPIB and NF-κB are the main molecular events driving cytarabine resistance development.Electronic supplementary materialThe online version of this article (10.1186/s12885-018-4346-1) contains supplementary material, which is available to authorized users.

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Protein phosphatase 2A regulates deoxycytidine kinase activity via Ser‐74 dephosphorylation

Cited by 6 publications

References 35 publications

The PPP2R1A cancer hotspot mutant p.R183W increases clofarabine resistance in uterine serous carcinoma cells by a gain-of-function mechanism

The PPP2R1A cancer hotspot mutant p.R183W increases clofarabine resistance in uterine serous carcinoma cells by a gain-of-function mechanism

dCK negatively regulates the NRF2/ARE axis and ROS production in pancreatic cancer

Bortezomib prevents cytarabine resistance in MCL, which is characterized by down-regulation of dCK and up-regulation of SPIB resulting in high NF-κB activity

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